Biology subjects

Goodrich-Blair, H.

Publications and source records attributed to Goodrich-Blair, H..

8 recordsLinked to original sources

bioRxiv2026

Development and behavior of Steinernema hermaphroditum nematodes are impacted by a bacterial flavin monooxygenase

Beneficial animal-microbe symbioses occur across the tree of life, with observable impacts on animal development and behavior. Some of these impacts occur when microbial enzymes act upon host-derived metabolites. The bacterial enzyme complex HpaBC hydroxylates aromatic compounds, including tyrosine and precursors of the neurotransmitter dopamine, raising the possibility that this enzyme may indirectly impact neurotransmitter-dependent phenotypes. The bacterium Xenorhabdus griffiniae, a beneficial intestinal symbiont of Steinernema hermaphroditum entomopathogenic nematodes, encodes HpaBC. Computational docking of X. griffiniae HpaB affinity for dopamine pathway substrates revealed energetically favorable docking of both dopamine and tyrosine, though neither was as favorable as the predicted affinity of HpaB for the canonical substrate, 4-hydroxyphenylacetate. We hypothesized that Xenorhabdus HpaBC may influence nematode host development or behavior through its potential action on tyrosine degradation or dopamine catabolism. In support of this idea, we demonstrate a negative correlation between X. griffiniae hpaBC expression and body length and egg-laying behavior in adult S. hermaphroditum nematodes. Moreover, hpaBC expression is necessary for X. griffiniae bacteria to colonize the nematode intestine in the infective juvenile stage. Taken together, these findings indicate that HpaBC may be a part of the metabolic crosstalk occurring between X. griffiniae and S. hermaphroditum at multiple stages of their shared life cycle. hpaBC orthologs are widely present across bacterial phyla including in pathogens and mutualists of plants and animals. Our findings raise the possibility that HpaBC could be a conserved mechanism by which host-associated bacteria influence physiology and behavior. ImportanceBeneficial animal-microbe interactions can shape development, physiology, and behavior in organisms across the tree of life, yet in many cases, the molecular mechanisms driving these interactions remain poorly understood. This study reveals that the bacterial metabolic enzyme complex HpaBC plays a role in the Steinernema-Xenorhabdus symbiosis, influencing nematode growth, reproduction, and host colonization. As hpaBC expression is conserved across many bacterial phyla, these findings highlight a potentially widespread mechanism for modulating animal physiology through microbial metabolism.

microbiology↗

bioRxiv2025

Intraspecific bacterial competition mediated by rapidly diversifying tailocin and prophage loci

Signatures of selection in microbial genomes are often linked to biotic interactions, notably resistance to host immunity or bacteriophage attack. Here, we highlight the importance of competitive interactions, specifically between conspecific bacteria, in shaping microbial genomes, using animal-associated Xenorhabdus bacteria. An aspect of microbial genome variation is the presence of diverse mobile genetic elements that distinguish bacterial genomes from their closely related kin. We found that compared to those across domain Bacteria, Xenorhabdus genomes contain among the highest proportion of phage-related genes, and that variation among strains in their total number of protein-coding genes is largely predicted by variation in total number of non-cargo phage genes per genome. A universal yet highly variable Xenorhabdus phage-related region encodes xenorhabdicin tailocins. This region ranged in length from 12 to 41 kilobases, and its specificity-determining main tail fiber varied from 341 to 1035 amino acids. Concomitant with this variation, tailocins produced by six strains of X. nematophila differed dramatically in particle length and killing profile. Intriguingly, while X. nematophila xenorhabdicins displayed common heterospecific killing activity, they varied in conspecific killing activity. We further demonstrate the ecological importance of xenorhabdicin diversity by associating intraspecific variation in killing profiles of mitomycin-induced lysates from 42 sympatric X. bovienii strains with genes from both xenorhabdicin-encoding loci and prophages. The susceptibility profiles of strains to lysates were associated with O-antigen biosynthesis genes. Overall, our data demonstrate that through bacteriophage-mediated genome diversification, an animal-associated bacterium can tailor its weaponry and defense systems to target the closest of relatives. IMPORTANCEMicrobes exist in complex communities with each other and with animal or plant hosts. Such biotic interactions can impose strong and variable selection on microbes and are predicted to foster diversity within lineages. By examining genomes from one bacterial genus, we show that phage-associated genes are responsible for an unusually high degree of variation in protein-coding gene content. Moreover, we link this variation to functional diversity in competitive interactions among Xenorhabdus strains. Different strains of Xenorhabdus bacteria frequently co-infect an insect, which simultaneously promotes strong selection for competitive dominance within an insect host as well as opportunities for horizontal gene flow. Additionally, genomic rearrangement and homologous recombination can provide phenotypic variation for selection. The tractability of Xenorhabdus for both lab and environmental studies make them powerful for understanding microbial genome evolution and competition and this study reveals the dominant role of bacteriophage and bacteriophage-like elements in these processes.

microbiology↗

bioRxiv2025

Genomes of the entomopathogenic nematode Steinernema hermaphroditum and its associated bacteria

As an entomopathogenic nematode (EPN), Steinernema hermaphroditum parasitizes insect hosts and harbors symbiotic Xenorhabdus griffinae bacteria. In contrast to other Steinernematids, S. hermaphroditum has hermaphroditic genetics, offering the experimental scope found in Caenorhabditis elegans. To enable biological analysis of S. hermaphroditum, we have assembled and analyzed its reference genome. This genome assembly has five chromosomal scaffolds and 83 unassigned scaffolds totaling 90.7 Mb, with 19,426 protein-coding genes having a BUSCO completeness of 88.0%. Its autosomes show higher densities of strongly conserved genes in their centers, as in C. elegans, but repetitive elements are evenly distributed along all chromosomes, rather than with higher arm densities as in C. elegans. Either when comparing protein motif frequencies between nematode species or when analyzing gene family expansions during nematode evolution, we observed two categories of genes preferentially associated with the origin of Steinernema or S. hermaphroditum: orthologs of venom genes in S. carpocapsae or S. feltiae; and some types of chemosensory G protein-coupled receptors, despite the tendency of parasitic nematodes to have reduced numbers of chemosensory genes. Three-quarters of venom orthologs occurred in gene clusters, with the larger clusters comprising functionally diverse pathogenicity islands rather than paralogous repeats of a single venom gene. While assembling the genome of S. hermaphroditum, we coassembled bacterial genomes, finding sequence data for not only the known symbiont, X. griffinae, but also for eight other bacterial genera. All eight genera have previously been observed to be associated with Steinernema species or the EPN Heterorhabditis, and may constitute a "second bacterial circle" of EPNs. The genome assemblies of S. hermaphroditum and its associated bacteria will enable use of these organisms as a model system for both entomopathogenicity and symbiosis.

genomics↗

bioRxiv2024

Bioinformatic discovery of type 11 secretion system (T11SS) cargo across the Proteobacteria

Type 11 secretion systems (T11SS) are broadly distributed among proteobacteria, with more than 3000 T11SS family outer membrane proteins (OMPs) comprising 10 major sequence similarity network (SSN) clusters. Of these, only 7, all from animal-associated cluster 1, have been experimentally verified as secretins of cargo, including adhesins, hemophores, and metal binding proteins. To identify novel cargo of a more diverse set of T11SS, we identified gene families co-occurring in gene neighborhoods with either cluster 1 or marine microbe-associated cluster 3 T11SS OMP genes. We developed bioinformatic controls to ensure perceived co-occurrences are specific to T11SS, and not general to OMPs. We found that both cluster 1 and cluster 3 T11SS OMPs frequently co-occur with single carbon metabolism and nucleotide synthesis pathways, but that only cluster 1 T11SS OMPs had significant co-occurrence with metal and heme pathways, as well as with mobile genetic islands, potentially indicating diversified function of this cluster. Cluster 1 T11SS co-occurrences included 2556 predicted cargo proteins, unified by the presence of a C-terminal {beta}-barrel domain, which fall into 141 predicted UniRef50 clusters and approximately 10 different architectures: 4 similar to known cargo and 6 uncharacterized types. We experimentally demonstrate T11SS-dependent secretion of an uncharacterized cargo type with homology to Plasmin sensitive protein (Pls). Unexpectedly, genes encoding marine cluster 3 T11SS OMPs only rarely co-occurred with the C-terminal {beta}-barrel domain and instead frequently co-occurred with DUF1194-containing genes. Overall, our results show that with sufficiently large-scale and controlled genomic data, T11SS-dependent cargo proteins can be accurately predicted. Impact StatementWe describe a novel method for controlling genomic neighborhood analyses and use this controlled co-occurrence technique to examine two distinct clusters of genes from the recently described type 11 secretion system (T11SS): Cluster 1, predominantly encoded by host-associated microbes and Cluster 3, encoded by marine microbes. We found that both clusters of predicted T11SS frequently co-occur with single carbon metabolism and nucleotide synthesis pathways, but only the host-associated cluster co-occurred with iron uptake pathways. Using these datasets, we predicted 2687 T11SS dependent cargo with approximately ten unique architectures, six of which have not previously been linked to T11SS. Finally, we validate our results by demonstrating T11SS-dependent secretion of a cargo protein with one of the novel architectures, plasmin-sensitive surface protein Pls from Haemophilus parahaemolyticus. Data SummaryThis paper describes genomic co-occurrences of T11SS OMP sequences originally identified through sequence similarity networking in a previous publication [3] as constituents of similarity clusters, 1 and 3. Accession IDs and assorted metadata for those clusters/networks are listed in Table S1 of that article: https://journals.asm.org/doi/suppl/10.1128/mbio.01956-21/suppl_file/mbio.01956-21-st001.xlsx. All other supporting data, accession IDs, code, and protocols have been provided within this article or through supplementary data files. Additionally, all supplemental data are available on Figshare (https://doi.org/10.6084/m9.figshare.28514069.v2) [1]. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=137 SRC="FIGDIR/small/615229v2_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@c9123dorg.highwire.dtl.DTLVardef@cc7295org.highwire.dtl.DTLVardef@76a60corg.highwire.dtl.DTLVardef@18f6c6d_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2024

Analyses of Xenorhabdus griffiniae genomes reveal two distinct sub-species that display intra-species variation due to prophages.

Nematodes of the genus Steinernema and their Xenorhabdus bacterial symbionts are lethal entomopathogens that are useful in the biocontrol of insect pests, as sources of diverse natural products, and as research models for mutualism and parasitism. Xenorhabdus play a central role in all aspects of the Steinernema lifecycle, and a deeper understanding of their genomes therefore has the potential to spur advances in each of these applications. Here, we report a comparative genomics analysis of Xenorhabdus griffiniae, including the symbiont of Steinernema hermaphroditum nematodes, for which genetic and genomic tools are being developed. We sequenced and assembled circularized genomes for three Xenorhabdus strains: HGB2511, ID10 and TH1. We then determined their relationships to other Xenorhabdus and delineated their species via phylogenomic analyses, concluding that HGB2511 and ID10 are Xenorhabdus griffiniae while TH1 is a novel species. These additions to the existing X. griffiniae landscape further allowed for the identification of two subspecies within the clade. Consistent with other Xenorhabdus, the analysed X. griffiniae genomes each encode a wide array of antimicrobials and virulence-related proteins. Comparative genomic analyses, including the creation of a pangenome, revealed that a large amount of the intraspecies variation in X. griffiniae is contained within the mobilome and attributable to prophage loci. In addition, CRISPR arrays, secondary metabolite potential and toxin genes all varied among strains within the X. griffiniae species. Our findings suggest that phage-related genes drive the genomic diversity in closely related Xenorhabdus symbionts, and that these may underlie some of the traits most associated with the lifestyle and survival of entomopathogenic nematodes and their bacteria: virulence and competition. This study establishes a broad knowledge base for further exploration of not only the relationships between X. griffiniae species and their nematode hosts but also the molecular mechanisms that underlie their entomopathogenic lifestyle.

microbiology↗

bioRxiv2023

Bacterial hemophilin homologs and their specific type eleven secretor proteins have conserved roles in heme capture and are diversifying as a family

Cellular life relies on enzymes that require metal cofactors, which must be acquired from extracellular sources. Bacteria utilize surface and secreted proteins to acquire such valuable nutrients from their environment. These include the cargo proteins of the type eleven secretion system (T11SS), which have been connected to host specificity, metal homeostasis, and nutritional immunity evasion. This Sec-dependent, Gram-negative secretion system is encoded by organisms throughout the phylum Proteobacteria, including human pathogens Neisseria meningitidis, Proteus mirabilis, Acinetobacter baumannii, and Haemophilus influenzae. Experimentally verified T11SS-dependent cargo include host metal acquisition proteins transferrin binding protein B (TbpB) and lactoferrin binding protein B (LbpB), as well as the hemophilin homologs heme receptor protein C (HrpC) and hemophilin A (HphA), the complement immune evasion protein factor-H binding protein (fHbp), and the host symbiosis factor nematode intestinal localization protein C (NilC). Secretion of each of these cargo proteins relies on a specific T11SS. Here, we examined the specificity of T11SS systems for their cognate cargo proteins using taxonomically distributed homolog pairs of T11SS and hemophilin cargo and explore the ligand binding ability of those hemophilin homologs. Our comparative ligand binding analysis of four hemophilin family proteins identified previously unknown ligand binding diversity within this protein family, which informed our description of structural features that are likely to contribute to heme/porphyrin binding specificity. In vivo expression of hemophilin homologs revealed that each was secreted in a specific manner by its cognate T11SS protein. Furthermore, secretion assays of chimeric hemophilin proteins revealed that specificity is predominantly dictated by the C-terminal domain of the cognate cargo. Meanwhile, the N-terminal effector domains of these T11SS-dependent cargo proteins feature porphyrin binding pockets that drive ligand binding affinity and specificity. In light of these results, we have termed this N-terminal domain the hemophilin ligand binding domain (Hlb) after its first characterized representative.

microbiology↗

bioRxiv2021

Chemical ecology of a tripartite symbiosis

Microbial symbiotic interactions, mediated by small molecule signaling, drive physiological processes of higher order systems. Metabolic analytic technologies advancements provide new avenues to examine how chemical ecology, or conversion of existing biomass to new forms, changes over a symbiotic lifecycle. We examine such processes using the tripartite relationship between nematode host Steinernema carpocapsae, its obligate mutualist bacterium, Xenorhabdus nematophila, and the insects they infect together. We integrate trophic, metabolomics, and gene regulation analyses to understand insect biomass conversion to nematode or bacterium biomass. Trophic analysis established bacteria as the primary insect consumers, with nematodes at trophic position 4.37, indicating consumption of bacteria and likely other nematodes. Significant, discrete metabolic phases were distinguishable from each other, indicating the insect chemical environment changes reproducibly during bioconversion. Tricarboxylic acid cycle components and amino acids were significantly affected throughout infection. These findings contribute to an ongoing understanding of how symbiont associations shape chemical environments. TeaserEntomopathogenic nematodes act as an apex predator in some ecosystems through altering chemical environments of their prey.

microbiology↗

bioRxiv2020

Beyond Slam: the DUF560 family outer membrane protein superfamily SPAM has distinct network subclusters that suggest a role in non-lipidated substrate transport and bacterial environmental adaptation

In host-associated bacteria, surface and secreted proteins mediate acquisition of nutrients, interactions with host cells, and specificity of tissue-localization. In Gram-negative bacteria, the mechanism by which many proteins cross or become tethered to the outer membrane remains unclear. The domain of unknown function (DUF)560 occurs in outer membrane proteins throughout Proteobacteria and has been implicated in host-bacteria interactions and lipoprotein surface exposure. We used sequence similarity networking to reveal three subfamilies of DUF560 homologs. One subfamily includes those DUF560 proteins experimentally characterized to date: NilB, a host-range determinant of the nematode-mutualist Xenorhabdus nematophila, and the surface lipoprotein assembly modulators Slam1 and Slam2, which facilitate msurface exposure of lipoproteins in Neisseria meningitidis (1, 2). We show that DUF560 proteins from a second subfamily facilitate secretion of soluble, non-lipidated proteins across the outer membrane. Using in silico analysis, we demonstrate that DUF560 gene complement correlates with bacterial environment at a macro level and host association at a species level. The DUF560 protein superfamily represents a newly characterized Gram-negative secretion system capable of lipoprotein surface exposure and soluble protein secretion with conserved roles in facilitating symbiosis. In light of these data, we propose that it be titled the type eleven secretion system (TXISS). ImportanceThe microbial constituents of a host associated microbiome are decided by a complex interplay of microbial colonization factors, host surface conditions, and host immunological responses. Filling such niches requires bacteria to encode an arsenal of surface and secreted proteins to effectively interact with the host and co-occurring microbes. Bioinformatic predictions of the localization and function of putative bacterial colonization factors are essential for assessing the potential of bacteria to engage in pathogenic, mutualistic, or commensal activities. This study uses publicly available genome sequence data, alongside experimental results from representative gene products from Xenorhabdus nematophila, to demonstrate a role for DUF560 family proteins in the secretion of bacterial effectors of host interactions. Our research delineates a broadly distributed family of proteins and enables more accurate predictions of the localization of colonization factors throughout Proteobacteria.

microbiology↗